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Ethanol Dose- and Time-dependently Increases α and β Subunits of Mitochondrial ATP Synthase of Cultured Neonatal Rat Cardiomyocytes

Keiko Mashimo¹, Peter G. Arthur² and Youkichi Ohno¹

¹Department of Legal Medicine, Nippon Medical School
²School of Biomedical, Biomolecular and Chemical Sciences, Faculty of Life and Physical Sciences, University of Western Australia, Australia

Mitochondria are target subcellular organelles of ethanol. In this study, the effects of ethanol on protein composition was examined with 2-dimensional electrophoresis of protein extracts from cultured neonatal rat cardiomyocytes exposed to 100 mM ethanol for 24 hours. A putative β subunit of mitochondrial ATP synthase was increased, which was confirmed by Western blot. The cellular protein abundances in the α and β subunits of ATP synthase increased in dose (0, 10, 50, and 100 mM)- and time (0.5 hour and 24 hours)-dependent manners. The DNA microarray analysis of total RNA extract demonstrated that gene expression of the corresponding messenger RNAs of these subunit proteins did not significantly alter due to 24-hour ethanol exposure. Therefore, protein expression of these nuclear-encoded mitochondrial proteins may be regulated at the translational, rather than the transcriptional, level. Alternatively, degradation of these subunit proteins might be decreased. Additionally, cellular ATP content of cardiomyocytes scarcely decreased following 24-hour exposure to any examined concentrations of ethanol. Previous studies, together with this study, have demonstrated that protein abundance of the α subunit or β subunit or both subunits of ATP synthase after ethanol exposure or dysfunctional conditions might differ according to tissue: significant increases in heart but decreases in liver and brain. Thus, it is suggested that the abundance of subunit proteins of mitochondrial ATP synthase in the ethanol-exposed heart, being different from that in the liver and brain, should increase dose-dependently through either translational upregulation or decreased degradation or both to maintain ATP production, as the heart requires much more energy than other tissues for continuing sustained contractions.

J Nippon Med Sch 2015; 82: 237-245

Keywords
ethanol, cardiomyocyte, ATP synthase, Western blot, DNA array

Correspondence to
Keiko Mashimo, Department of Legal Medicine, Nippon Medical School, 1-1-5 Sendagi, Bunkyo-ku, Tokyo 113-8602, Japan
kei-mam@nms.ac.jp

Received, July 23, 2015
Accepted, October 6, 2015